The mRNAexpression study on small amount of Theileria annulata lymph node biopsy sample using SMART-cDNA technology
2013
Sadr Shirazi, Nastaran | Shayan, Parviz | Eckert, Berigiteh | Ebrahimzadeh, Elaheh | Jafari, Sedigheh
BACKGROUND: A major issue in many gene expressionstudies utilizing small amount of biological materials is thelimited quantity of RNApurified from clinical samples, which isoften used for RT-PCR or standard Northern blot analysis.OBJECTIVES: The SMART cDNA synthesis method and subsequentSMART-cDNA-PCR technique was used to analyse 3genes in macroschizonts of Theileria annulata in small lymphnode biopsy material. METHODS: The SMART-cDNA of TaSpgene was cloned in pTZ57R/T-vector and sequenced. We focusedon genes encoding surface proteins TaSp, TaD and HSP70.RESULTS: Our results showed that SMART cDNA dependablyreproduces the expression profile found in messenger RNA. TheRT-SMART-PCR showed the amplification of the processedmRNAs. The sequencing analysis showed that the amplifiedcDNA was coded for TaSp protein in Theileria annulata.CONCLUSIONS: It was concluded that the SMART PCRtechnique is practical for amplification of complete sequence ofmRNAs in the form of cDNAs, and therefore for gene expressionstudies if only small amounts of starting material are available.
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